马兜铃酸诱导大鼠肾小管上皮细胞表型转化和胶原累积及垂盆草提取物的作用  

Sedum sarmentosum Bunge extract inhibits epithelial-mesenchymal transition and collagen accumulation in aristolochic acid-treated rat renal tubular epithelial cells

作　　者：陆红[1] 胡丽萍[1] 洪丹[2] 洪炜龙[3] 林成成[3] 王斯璐[3] 陈必成[3] 白永恒[3] 

机构地区：[1]温州医科大学附属第一医院医学检验中心,浙江温州325000 [2]温州医科大学生命科学学院,浙江温州325035 [3]温州医科大学附属第一医院外科实验室,浙江温州325000

出　　处：《中国病理生理杂志》2013年第12期2172-2178,共7页Chinese Journal of Pathophysiology

基　　金：浙江省自然科学基金资助项目(No.LQ12H050001;No.LY12H050004);温州市科技计划(No.Y20110028;No.Y20110072)

摘　　要：目的:研究垂盆草(SSB)提取物对马兜铃酸(AA)诱导的肾小管上皮细胞表型转化和胶原累积的作用,并初步探讨可能的分子机制。方法:将体外培养的大鼠肾小管上皮细胞(NRK-52E)分为:(1)溶剂对照组:未加入SSB和AA;(2)AA损伤组:只加AA,浓度为1~100 mg/L;(3)SSB提取物干预组:在加入10 mg/L AA基础上,同时加入SSB提取物(10~2 000 mg/L)。细胞培养24 h后,酶联免疫吸附试验检测转化生长因子β1(TGF-β1)含量;细胞免疫荧光染色检测肌成纤维细胞标志物α-平滑肌肌动蛋白(α-SMA)、上皮细胞标志物E-cadherin和基质成分III型胶原的表达;实时荧光定量PCR检测α-SMA、E-cadherin、骨形成蛋白7(BMP-7)和I型胶原mRNA的表达。结果:AA可诱导NRK-52E细胞呈现纤维化样改变;1 mg/L和10 mg/L AA不仅可增加基质成分I型和III型胶原的表达量,同时也可促进α-SMA表达,抑制E-cadherin的表达。用SSB提取物干预后,AA所致的纤维化改变明显减轻;SSB提取物下调了α-SMA、I型和III型胶原的表达,并且促进E-cadherin和BMP-7的表达。此外,SSB提取物也抑制TGF-β1的分泌,并呈浓度依赖性。结论:应用SSB提取物干预可明显降低AA所致的肾纤维化效应,可能的机制为SSB提取物降低TGF-β1的表达,抑制小管上皮细胞的表型转化,进而抑制胶原的累积。AIM： To investigate the effect of Sedum sarmentosum Bunge （SSB） extract on epithelial-mesenchy- mal transition （EMT） and collagen accumulation induced by aristolochic acid （AA） in renal tubular epithelial cells. METHODS： Rat renal tubular epithelial NRK-52E cells were randomly divided into 3 groups, including control group （only treated with solvent）, AA group （treated with AA at concentrations ranging from 1 to 100 mg/L） and SSB group （treated with AA at a concentration of 10 mg/L plus SSB extract at concentrations ranging from 10 to 2 000 mg/L）. After cultured for 24 h, the morphology of the NRK-52E cells was observed under inverted phase-contrast microscope. The level of transforming growth factor 13~ （TGF-~I） in the culture supernatant was measured by ELISA. Immunofluorescent analysis was performed to detect the expression of epithelial marker c^-smooth muscle actin （ ct-SMA）, mesenchymal marker E-cadherin, and extracellu- lar cell matrix component type III collagen. The mRNA expression of E-cadherin, c^-SMA, bone morphogenetic protein 7 （BMP-7） and type I collagen was also quantified by real-time PCR. RESULTS： Fibrosis-like reaction observed under mi- croscope was obviously increased in AA-treated NRK-52E cells, and aggravated as the increase in the concentration of AA. AA at concentrations of 1 and 10 mg/L increased the expression of α-SMA, type I and type III collagens, and decreased the expression of E-cadherin. With SSB extract treatment, fibrosis in NRK-52E cells was alleviated, accompanied with the decreasing expression of α-SMA, type I and type III collagen, and the enhancing expression of E-cadherin and BMP-7. Mo-reover, SSB extract down-regulated TGF-β level in a concentration-dependent manner. CONCLUSION： AA-induced fi- brosis-like reaction in renal tubular epithelial cells is reduced by the treatment with SSB extract. The possible mechanism is that SSB extract decreases TGF-β1 level, and inhibits renal EMT and collagen accumulation induced by AA.

关 键 词：垂盆草 马兜铃酸 转化生长因子β1 上皮-间充质转化 胶原 

分 类 号：R285.5[医药卫生—中药学]